Immunological memory is the fundamental goal of vaccine development. Therefore, it is imperative to understand the molecular pathways and gene networks that are involved in the generation of memory responses to infection/vaccination. Recent studies have shown that vaccines geared towards T cell immunity may provide better protection. Over the past few years, the cellular and molecular mechanisms behind memory CD8 T cell development have been elucidated. The generation of CD8 T cell memory is a highly orchestrated, multistep process that can be modulated by many factors. The cellular composition at the site of priming and soluble factors in a specific microenvironment can exert profound effects on the outcome of CD8 T cell responses. Therefore, it is important to understand the different T cell intrinsic and extrinsic factors that participate in the CD8 T cell differentiation and memory generation. In our preliminary studies, we observed that route of infection has a profound effect on CD8 T cell memory generation. Infection by either subcutaneous or intravenous routes generated comparable primary CD8 T cell responses to both bacterial and viral infections but subcutaneous route of infection failed to generate CD8 T cell memory. These results suggest that the route of infection/immunization can a have profound effect on the outcome of CD8 T cell responses and the mechanisms that regulate such differential responses need further investigation. Therefore, in the present proposal we precisely aim to characterize the role of resident DCs and cytokine milieu in the specific priming microenvironments in the differential programming of CD8 T cell responses. Consistent with the lack of generation of long term CD8 T cell memory we discovered differential gene expression profiles in CD8 T cells primed in the spleen versus the draining lymph nodes. We find that CD8 T cells that are primed in the spleen but not in the lymph nodes express genes that have been previously implicated in the development of CD8 memory T cells. Our overall goals of this proposal are to understand the cellular and molecular mechanisms by which the route of infection determines the outcome of CD8 T cell immunity. In specific aim 1, we will study the role of specific dendritic cells in the spleen and lymph nodes and understand if they play a role in determining the fate of activated CD8 T cells. In specific aim 2, we will investigate the molecular mechanisms by which CD8 T cell memory formation is regulated by certain genes we have discovered in our preliminary work. These studies will be critical to understand how different routes of immunization of infections can influence priming and development of memory and the findings will be very useful for developing strategies to induce protective immunity against pathogens.